Label-free biosensing with singular-phase-enhanced lateral position shift based on atomically thin plasmonic nanomaterials

Rapid plasmonic biosensing has attracted wide attention in early disease diagnosis and molecular biology research. However, it was still challenging for conventional angle-interrogating plasmonic sensors to obtain higher sensitivity without secondary amplifying labels such as plasmonic nanoparticles. To address this issue, we developed a plasmonic biosensor based on the enhanced lateral position shift by phase singularity. Such singularity presents as a sudden phase retardation at the dark point of reflection from resonating plasmonic substrate, leading to a giant position shift on reflected beam. Herein, for the first time, the atomically thin layer of Ge2Sb2Te5 (GST) on silver nanofilm was demonstrated as a novel phase-response-enhancing plasmonic material. The GST layer was not only precisely engineered to singularize phase change but also served as a protective layer for active silver nanofilm. This new configuration has achieved a record-breaking largest position shift of 439.3 μm measured in calibration experiments with an ultra-high sensitivity of 1.72 × 108 nm RIU−1 (refractive index unit). The detection limit was determined to be 6.97 × 10−7 RIU with a 0.12 μm position resolution. Besides, a large figure of merit (FOM) of 4.54 × 1011 μm (RIU∙°)−1 was evaluated for such position shift interrogation, enabling the labelfree detection of trace amounts of biomolecules. In targeted biosensing experiments, the optimized sensor has successfully detected small cytokine biomarkers (TNF-α and IL-6) with the lowest concentration of 1 × 10−16 M. These two molecules are the key proinflammatory cancer markers in clinical diagnosis, which cannot be directly screened by current clinical techniques. To further validate the selectivity of our sensing systems, we also measured the affinity of integrin binding to arginylglycylaspartic acid (RGD) peptide (a key protein interaction in cell adhesion) with different Mn2+ ion concentrations, ranging from 1 nM to 1 mM.

measured by the tapping mode of Atomic Force Microscope (Nanoscope, Bruker Inc., France).The mean surface roughness (Ra) in this region is 0.70 nm.Fig. S1c provides a wider optical surface profile for characterizing the thickness of GST capping layer, measured by Digital Holography Microscope (R2100, Lyncée Tec Inc., France).The optical profile is the optical path length distribution (OPL) on surface, which is determined by  ×   . represent the real surface profile while   denotes the material refractive index at the microscope working wavelength (675nm).Fig. S1d is a cross-section view of the optical profile illustrating the GST layer thickness measurement.The mean OPL difference between the top surface of GST capping and surround silver ranges from 3.53 to 4.10 nm over 10 profiles.The GST refractive index at 675nm is 3.93 S1 .Therefore, the thickness of GST layer should be 0.90 to 1.04 nm.

Supplementary note 2: Substrate stability in solution
An immersion test in PBS was applied to validate the stability of bare silver substrate and the silver substrate with 1nm GST capping layer.In Figs.S2a and S2b, the GST capped region did not show observable degeneration after 1 hour and 24 hours of immersion, while the bare silver region was visibly prone to oxidation darkening with dense black spots (Fig. S2b).Moreover, Figs.S2c and S2d provide the reflectance decay after the immersion test to quantitively evaluate the degeneration on substrates.The decay curves were calculated by subtracting the after-immersion spectrum to raw spectrum.After 24 hours of immersion, the reflectance on silver substrate has reduced by 60%.In contrast, there was no observable decay on silver-GST substrate.Such enhanced stability of sliver-GST substrate can ensure the sensor performance during the measurement as the time for a single test (Fig. 6, 7) is around half an hour.The relationship between reflectance and phase is illustrated in Fig. S5a.The phase changing rate will be accelerated if the reflection is suppressed.At the minimal reflection point, the phase changing rate is maximized, resulting in an abrupt phase jump.Such abrupt phase change can be topologically explained on a complex plane as depicted in Fig. S5b.The X and Y axes of this plane respectively represent the real and image parts of complex reflection coefficient   .  as the norm of   denotes the reflectance.Φ as the phase angle of   denotes the phase of incident light.If   vector is approaching the origin on complex plane, i.e.,   → 0 , the change rate of phase angle will be accelerated that eventually results in an abrupt jump.Furthermore, according to equation S1: ), the sharpened phase jump brings a larger GH shift and higher sensitivity.Therefore, the suppressed reflection is the direct factor leading to better GH sensing performance.
The 1nm GST layer was employed to further minimize the reflectance at the resonance angle of the sensing device.Since GST alloy is an absorptive material in visible to NIR range S2,S3 .As given in Figs.S5c and S5d, such GST layer thickness is designed to avoid the over flatten of reflectance curve and subsequent mismatching of phase singularity condition.Also, it is not possible to use metallic layer only to achieve this maximized light absorption.We plotted the silver substrate response by tuning the thickness as shown in Fig. S5c.The minimum reflectance is 2.69×10 -4 , not as low as the result achieved by silver-GST substrate (1.03×10 -6 )

Table S1: Summary table of sensing performance related parameters between different substrates and operating wavelengths
As shown in Fig. S6, in order to do a better comparison between silver substrates and conventional gold substrates.we have fabricated the gold sensing substrates with similar thickness parameters.In Figs S6a and S6b, the reflectance matching results indicate that the fabricated substrates have 38nm Au and 1nm GST.As we can see for both 785 nm and 633 nm, the GH width curve and the experimentally measured sensitivity all have a similar trend as for the silver substrates.For example, in Figs.S6c and S6d, the maximum GH shift of 56.6 μm were found on GST coated substrate with 785 nm excitation as well.However, compared to the silver-GST substrate operated at same wavelength, the decreased peak value of GH shift on gold-GST substrate resulted in nearly 10 times lower experimental sensitivity as shown in Figs.S6e, S6f and 4c, 4d.
The sensing performance related parameters are listed in Tabel S1, for the comparison between different sensing substrates and sensing conditions.To quantitively compare the GH sensing response, the figure of merit (FOM) defined by sensitivity/FWHM was introduced in this table with the unit of μm (RIU•°) -1 .The gold-GST sensing substrate operated at 785 nm has a FOM value of 1.24×10 5 , which was much lower than that of silver-GST substrate (4.54×10 11).(5) Where for non-magnetic substance: No need for signal amplification with contrast agents (label-free) Bright field micrographs of bare silver and 1nm GST coated silver substrate after 1-(a) and 24-hours (b) immersion in PBS buffer.The pictures are taken by digital microscope.c, d Reflectance decay measured on (c) GST coated silver (d) bare silver substrate after immersion test.The decay is measured by spectrometer in comparing with the spectrum acquired before immersion.Zoom in subplots inside illustrate the reflectance decay around the operating wavelength of 785nm.The negative value shown in (c) is due to the enlarged reflectance after buffer immersion and rinse.The spectrum shown in Figs S2c and S2d have a sudden drift at 700nm.It results from the misalignment of the dual sensors in spectrometer and independent to reflectance measurement.